The lac operon has served as the paradigm to study gene regulation. The x-ray crystallographic analyses of the lac repressor and its complexes with effector molecules and lac operator DNA will provide the structural platform for understanding gene regulation. Our primary interest is to understand how the repressor recognizes the specific sequence of the operator DNA. It is also important to establish the binding sites for inducers and anti-inducers, and then, to establish how these effectors control DNA binding. These structures will also provide information about the structural aspects of allostery. Genetic data has delineated four functional regions of the repressor. Specific point mutants have been isolated with phenotypes that are defective in: operator DNA binding, inducer binding, signal transmission, and quaternary associations. Our main focus will be to determine the 3-dimensional structure of the lac repressor, to experimentally elucidate the structure the effectors binding, and determine the structure of cocrystals of the lac repressor with a symmetric 21 base pair lac operator. The structure of the lac repressor will allow us to address such questions as (1) how does the repressor recognize specific sites on the DNA, (2) how do inducers and anti-inducers affect the conformation of the repressor, and ultimately (3) how does the structure relate to the allosteric mechanism used to regulate gene expression. Over four thousand single amino acid substitutions have been analyzed for phenotype. As a consequence, there have been more functional variants produced by amino acid substitutions in the lac repressor than in any other protein including hemoglobin. The site directed mutagenesis on this project is the most comprehensive of all proteins. An analysis of the mutants with respect to the tertiary and quaternary structures that will be the result of this project will provide exciting insights into protein folding. With the three dimensional structure well in hand, the lac repressor will be the most complete system to study protein folding. The structural studies of the native lac repressor, the repressor-inducer complex and cocrystals of the repressor-DNA complex will provide a long awaited structural framework to describe more completely the lac operon - - the prototypical gene regulation system.